The following abbreviations are herewith defined, at least some of which are referred to within the following description of the state-of-the-art and the present invention. The reference to the “present invention” or “invention” used herein relates to exemplary embodiments and not necessarily to every embodiment that is encompassed by the appended claims.    ADME-Tox Absorption, Distribution, Metabolism, Excretion and Toxicity    ASGPR Asialoglycoprotein Receptor    ECM Extracellular Matrices    US FDA United States Food Drug Administration    HTS High-Throughput Screening    LC Liquid Chromatography    MS Mass Spectroscopy    NCE New Chemical Entity    SBS Society of Biomolecular Screening    TEER Trans Epithelial Electrical Resistance
The following references are cited below in the description of the state of-the-art, where their contents are hereby incorporated by reference herein.    1. Yau Yi Lau et al. “Evaluation of a Novel In Vitro Caco-2 Hepatocyte Hybrid System for Predicting In Vivo Oral Bioavailability” Drug Metabolism and Disposition, Vol. 32, No. 9, pp. 937-942, 2004.    2. “Transwell® Permeable Supports: Including Snapwell™ and Netwell™ Inserts—Instructions for Use” Corning Inc., September 2007.    3. Sue Hyung Choi et al. “Feasibility of Simple Double-Layered Coculture System Incorporating Metabolic Processes of the Intestine and Liver Tissue: Application to the Analysis of Benzo[a]pyrene Toxicity”, Toxicology in Vitro, vol. 18, pages 393-402, 2004.    4. U.S. Department of Health and Human Services et al. “Guidance for Industry: Drug Interaction Studies-Study, Design, Data Analysis, and Implications for Dosing and Labeling”, Clinical Pharmacology, September 2006.
In the search for new therapeutic drugs, pharmaceutical companies utilize many different methods to determine whether a compound or molecule known as a NCE has a desired biological activity. These methods often entail examining the ADME-Tox of the NCE, as well as determining the NCE's level of effectiveness for the targeted therapeutic indication including pharmacokinetic parameters. One type of assessment examines the “first pass effect”. This assessment involves experimental determination of the bioavailabilty of the NCE following its absorption through the digestive tract and then its metabolism by the liver. Commonly, the assessment of the “first pass effect” requires two separate in vitro assays to be conducted, and the data combined, to determine the intestinal permeability and the hepatic metabolism. If desired, additional studies may be conducted to determine target selectivity, efficacy and dosage (reference no. 1).
A well-known method used today to examine the intestinal absorption of a NCE is known as the Caco 2 cell-based assay which is typically conducted on permeable supports such as the ones sold under the brand name of Transwell™ and manufactured by Corning Inc. (reference no. 2). The design of the Transwell™ permeable support facilitates the development of Caco 2 cell polarization to create more in vivo-like test conditions. Researchers from the Schering-Plough Research Institute have expanded the utility of the Caco 2 cell-based assay by adding hepatocytes in the nutrient medium to a Transwell™ receiver plate which receives the Transwell™ permeable support. In this way, the researchers were able to more accurately predict the oral bioavailability of NCE's. However, the hepatic cell viability under these conditions during a 3 hour incubation period was only 50-70%, limiting the potential of this method (reference no. 1). Another group of researchers from the University of Tokyo co-cultured Caco-2 cells on the Transwell™ permeable support with monolayers of Hep G2 cells growing on the inner surface of the Transwell™ receiver plate. While useful for some assays, the Hep G2 cells did not maintain the functions that are representative of in vivo hepatocytes (reference no. 3).
The current US FDA Guidance regarding drug interaction studies like the first pass assay recommends the use of in vitro assays with fresh or cryopreserved human hepatocytes due to species specific responses (reference no. 4). However, it is well known that primary hepatocytes loose differentiated function rapidly in standard cell culture conditions on tissue culture treated polystyrene. The loss of normal differentiated hepatocyte function decreases the in vivo-like conditions and hence also decreases the relevance of experimental data in ADME-Tox and pharmacokinetic in vitro assays. Thus, any enhancement of test equipment and conditions to provide a more in vivo-like situation for the cells in culture would greatly improve the relevance of the information which is gained by these types of experiments.